To study the flexural fatigue performance of prestressed reactive powder concrete (RPC)-normal concrete (NC) composite beams, four identical reduced-scale model beams were designed and produced on the basis of a 32 m span T-girder of Chinese railway. One of the beams suffered static load and others were under bending fatigue test. Strain distribution in normal section, strain variation in compressed NC, crack propagation, and the development of fatigue deflection and stiffness were analyzed. Furthermore, flexural property of the composite beams without failure after fatigue loading were compared with the composite beams that did not experience fatigue loading and the NC beams with identical structural size and reinforcement condition. Results show that fatigue failure of prestressed RPC-NC composite beams was caused by the fatigue fracture of non-prestressed longitudinal bars, which was the same as balanced-reinforced NC beam. Strain in normal section was always approximately linear with the height of the section under fatigue load, which means the plane section assumption can be applied to prestressed RPC-NC composite beams. Flexural ductility of the composite beams without failure after fatigue loading declined compared with the composite beam before fatigue loading, while it was still greater than NC beam. Stiffness degradation formula of test beams related to fatigue loading cycles was obtained by fitting experimental data, which can be a reference for the design of prestressed RPC-NC composite beams.